1. Volume 10, Issue 7, Pages 891-899 (July 2002)
Crystal Structure of the FHA Domain of the Chfr Mitotic Checkpoint Protein and Its Complex with Tungstate 
Elena S. Stavridi, Yentram Huyen, Ivy R. Loreto, Daniel M. Scolnick, Thanos D. Halazonetis, Nikola P. Pavletich, Philip D. Jeffrey 
Structure 
Volume 10, Issue 7, Pages (July 2002)
DOI: /S (02)

2. Figure 1 Structure of the Chfr FHA Domain
(A) Structure of the segment-swapped dimer. The two polypeptide chains are colored red and green, respectively. N, N terminus; C, C terminus.
(B) Comparison of the structures of the Chfr FHA and Rad53 N-terminal FHA domains. Only one domain of Chfr is shown, and the β strands involved in segment swapping are not shown in their entirety. In both structures, strands 1–7 and 8–11 are colored green and red, respectively, to maintain the same coloring scheme as in (A).
Structure  , DOI: ( /S (02) )

3. Figure 2 Structure of Conserved Regions
(A) Amino acid sequence and secondary structure of the crystallized Chfr FHA domain (residues 14–124) aligned to conserved sequences (boxes A–D) of FHA domains from other proteins. The amino acid sequence numbering refers to the full-length proteins and indicates the position of the conserved boxes or, for Chfr, marks the entire sequence and selected residues. Rad53-N and Rad53-C, N- and C-terminal FHA domains of budding yeast Rad53; Dun1, FHA domain of budding yeast Dun1 checkpoint kinase; NBS1, FHA domain of human DNA repair and cell cycle checkpoint protein NBS1; MNF, FHA domain of mouse MNF transcription factor; Fkh1, FHA domain of budding yeast Fkh1 transcription factor.
(B–C) Comparison of the structures of Chfr FHA and Rad53 N-FHA domains showing the side chains of amino acids conserved in FHA domains. The Rad53 N-FHA domain is bound to a phosphothreonine (pT)-containing peptide. Backbone of the FHA domains corresponding to boxes A–D, orange; phosphopeptide, light blue; phosphate group, red. D+3, aspartic acid three residues C-terminal to the phosphothreonine.
Structure  , DOI: ( /S (02) )

4. Figure 3
Structure of the Chfr-Tungstate Complex and Comparison to Rad53 FHA Domain-Phosphopeptide Structures
(A–D) Comparison of the structures of native Chfr, the Chfr-tungstate complex, and the Rad53 N-FHA and C-FHA domains bound to phosphothreonine (pT)- and phosphotyrosine (pY)-containing peptides, respectively. Backbone of the FHA domains corresponding to boxes A–D, orange; phosphopeptides, light blue; phosphate and tungstate (WO4) groups, red. D+3, aspartic acid three residues C-terminal to the phosphothreonine.
(E) Superimposition of the structures of the Chfr-tungstate and Rad53 N-FHA and C-FHA domain-phosphopeptide complexes. FHA domain backbones, gray (various shades); side chains that contact tungstate/phosphate, red (Chfr), green (Rad53 N-FHA), and blue (Rad53 C-FHA); tungstate ion that binds to Chfr, red; phosphothreonine (pT) residue that contacts the Rad53 N-FHA domain, yellow and green; phosphotyrosine (pY) residue that contacts the Rad53 C-FHA domain, yellow and blue.
(F) Alignment of Chfr FHA and Rad53 N-FHA and C-FHA domain residues involved in tungstate/phosphate binding. Residues whose side chains are within 3.2 Å of a tungstate/phosphate oxygen atom, red; residues whose backbone nitrogen (N) contacts a tungstate/phosphate oxygen atom, blue.
Structure  , DOI: ( /S (02) )

5. Figure 4 Segment Swapping
(A) Refined monomer and segment-swapped dimer models in the P3(2)21 space group. The hydrogens of the Lys86 side chain are shown to indicate the presence of steric hindrance in the monomer model. The two polypeptide chains are colored green and red, respectively.
(B) Simulated annealing omit electron density maps contoured at 1.5 σ. The maps were generated using the refined monomer model in the P3(2)21 space group (left) or the refined segment-swapped dimer model in the P2(1)2(1)2(1) space group (right), with all atoms within 3 Å of Lys86 omitted. For both maps, the refined segment-swapped dimer models are shown to indicate that these models fit the omit electron density maps.
(C) Analytical ultracentrifugation analysis of the polypeptide containing residues 14–128 of Chfr. Lower panel: protein concentrations at equilibrium for the three initial loading concentrations (1.0, 0.5, and 0.25 mg/ml) at 33,800 rpm and 4°C. Circles, raw data; lines, fitted curves for monomer-dimer equilibrium. Upper panels: residuals of the fitted curves to the data points for each concentration from highest to lowest (top to bottom, respectively).
(D) Gel filtration chromatography elution profiles of Chfr FHA domain immediately after purification (Stock) or after extraction from amorphous precipitate (Precip.) or crystals (Crystal). The Stock, Precip., and Crystal elution profiles correspond to molecular sizes of 11.6, 11.5, and 20.6 kDa, respectively.
Structure  , DOI: ( /S (02) )